title: Identification of an enhancer that increases miR-200b~200a~429 gene expression in breast cancer cells.
abstract: The miR-200b~200a~429 gene cluster is a key regulator of EMT and cancer metastasis, however the transcription-based mechanisms controlling its expression during this process are not well understood. We have analyzed the miR-200b~200a~429 locus for epigenetic modifications in breast epithelial and mesenchymal cell lines using chromatin immunoprecipitation assays and DNA methylation analysis. We discovered a novel enhancer located approximately 5.1kb upstream of the miR-200b~200a~429 transcriptional start site. This region was associated with the active enhancer chromatin signature comprising H3K4me1, H3K27ac, RNA polymerase II and CpG dinucleotide hypomethylation. Luciferase reporter assays revealed the upstream enhancer stimulated the transcription of the miR-200b~200a~429 minimal promoter region approximately 27-fold in breast epithelial cells. Furthermore, we found that a region of the enhancer was transcribed, producing a short, GC-rich, mainly nuclear, non-polyadenylated RNA transcript designated miR-200b eRNA. Over-expression of miR-200b eRNA had little effect on miR-200b~200a~429 promoter activity and its production did not correlate with miR-200b~200a~429 gene expression. While additional investigations of miR-200b eRNA function will be necessary, it is possible that miR-200b eRNA may be involved in the regulation of miR-200b~200a~429 gene expression and silencing. Taken together, these findings reveal the presence of a novel enhancer, which contributes to miR-200b~200a~429 transcriptional regulation in epithelial cells.
authors.affiliation: Centre for Cancer Biology, SA Pathology, Adelaide, South Australia, Australia ; School of Molecular and Biomedical Science, the University of Adelaide, Adelaide, South Australia, Australia.
authors.forename: Joanne L
authors.initials: JL
authors.lastname: Attema
authors.forename: Andrew G
authors.initials: AG
authors.lastname: Bert
authors.forename: Yat-Yuen
authors.initials: YY
authors.lastname: Lim
authors.forename: Natasha
authors.initials: N
authors.lastname: Kolesnikoff
authors.forename: David M
authors.initials: DM
authors.lastname: Lawrence
authors.forename: Katherine A
authors.initials: KA
authors.lastname: Pillman
authors.forename: Eric
authors.initials: E
authors.lastname: Smith
authors.forename: Paul A
authors.initials: PA
authors.lastname: Drew
authors.forename: Yeesim
authors.initials: Y
authors.lastname: Khew-Goodall
authors.forename: Frances
authors.initials: F
authors.lastname: Shannon
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Goodall
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics*; D045744:Cell Line, Tumor; D002843:Chromatin / Q000235:genetics; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D057890:Epigenomics / Q000379:methods; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012313:RNA / Q000235:genetics; D024363:Transcription Initiation Site
chemical_list: D002843:Chromatin; D035683:MicroRNAs; D012313:RNA